A large molecular weight (Mr 300,000) form of phosphoprotein phosphatase was extracted from partially delipidated human CNS myelin. On ethanol treatment this phosphatase converted to a small Mr species which was then purified to apparent homogeneity by chromatography over DEAE-cellulose and Sephadex G-100 columns. The Mr of the smaller phosphatase was estimated at 46,000 by sodium dodecyl sulfate polyacrylamide gel electrophoresis and by chromatography over a calibrated Sephadex G-100 column. For optimal activity the enzyme required Mn2 ion and dithiothreitol. The purified phosphatase did not cleave p-nitrophenyl phosphate.